Adiponectin and insulin in grey seals during suckling and fasting: relationship with nutritional state and body mass during nursing in mothers and pups

Animals that fast during breeding and/or development, such as phocids, must regulate energy balance carefully to maximize reproductive fitness and survival probability. Adiponectin, produced by adipose tissue, contributes to metabolic regulation by modulating sensitivity to insulin, increasing fatty acid oxidation by liver and muscle, and promoting adipogenesis and lipid storage in fat tissue. We tested the hypotheses that (1) circulating adiponectin, insulin, or relative adiponectin gene expression is related to nutritional state, body mass, and mass gain in wild gray seal pups; (2) plasma adiponectin or insulin is related to maternal lactation duration, body mass, percentage milk fat, or free fatty acid (FFA) concentration; and (3) plasma adiponectin and insulin are correlated with circulating FFA in females and pups. In pups, plasma adiponectin decreased during suckling (linear mixed-effects model [LME]: T = 4.49; P < 0.001) and the early postweaning fast (LME: T = 3.39; P = 0.004). In contrast, their blubber adiponectin gene expression was higher during the early postweaning fast than early in suckling (LME: T = 2.11; P = 0.046). Insulin levels were significantly higher in early (LME: T = 3.52; P = 0.004) and late (LME: T = 6.99; P < 0.001) suckling than in fasting and, given the effect of nutritional state, were also positively related to body mass (LME: T = 3.58; P = 0.004). Adiponectin and insulin levels did not change during lactation and were unrelated to milk FFA or percentage milk fat in adult females. Our data suggest that adiponectin, in conjunction with insulin, may facilitate fat storage in seals and is likely to be particularly important in the development of blubber reserves in pups.

HSP70 abundance and antioxidant capacity in feeding and fasting gray seal pups: suckling is associated with higher levels of key cellular defenses

Heat shock proteins (HSPs) and antioxidants are key cellular defenses against stress. Seals routinely undergo protracted fasting, which is normally associated with physiological stress in other animals. We tested the hypotheses that (1) relative HSP70 protein abundance is higher in liver and blubber of fasting relative to suckling wild gray seal pups; (2) differences in HSP70 are mirrored in tissue superoxide dismutase (SOD) and catalase activity, as well as glutathione levels; (3) extracellular HSP70 correlates with hepatic and blubber HSP70 abundance; and (4) protein carbonylation, an index of oxidative damage, is lower in tissues with higher levels of these cellular stress markers. In contrast to our expectation, suckling pups had higher relative HSP70 abundance and glutathione levels in liver and blubber and higher hepatic catalase activity. Plasma HSP70 did not correlate with liver or blubber abundance of the protein. Suckling pups did not experience greater protein carbonylation, suggesting that cellular protective mechanisms prevent protein damage despite an apparent increase in cellular stress. SOD activity was not affected by nutritional state, but in blubber tissue, it was positively correlated with blubber thickness. Greater requirements for antioxidants and HSPs in suckling pups or in animals with thicker blubber could arise from rapid protein synthesis, high metabolic fuel availability, and/or exposure to lipophilic toxins. Developmental and nutritional changes in cellular defenses have important implications for gray seals’ susceptibility to additional stress exposure.

Liver glucose-6-phosphatase proteins in suckling and weaned grey seal pups: structural similarities to other mammals and relationship to nutrition, insulin signalling and metabolite levels

Phocid seals have been proposed as models for diabetes because they exhibit limited insulin response to glucose, high blood glucose and increasing insulin resistance when fasting. Liver glucose-6-phosphatase (G6Pase) catalyses the final step in glucose production and is central to glucose regulation in other animals. G6Pase comprises a translocase (SLC37A4) and a catalytic subunit (G6PC). G6PC and SLC37A4 expression and activity are normally regulated by nutritional state and glucostatic hormones, particularly insulin, and are elevated in diabetes. We tested the hypotheses that (1) grey seal G6PC and SLC37A4 cDNA and predicted protein sequences differ from other species’ at functional sites, (2) relative G6Pase protein abundances are lower during feeding than fasting and (3) relative G6Pase protein abundances are related to insulin, insulin receptor phosphorylation and key metabolite levels. We show that G6PC and partial SLC37A4 cDNA sequences encode proteins sharing 82–95 % identity with other mammals. Seal G6PC contained no differences in sites responsible for activity, stability or subcellular location. Several substitutions in seal SLC37A4 were predicted to be tolerated with low probability, which could affect glucose production. Suckling pups had higher relative abundance of both subunits than healthy, postweaned fasting pups. Furthermore, relative G6PC abundance was negatively related to glucose levels. These findings contrast markedly with the response of relative hepatic G6Pase abundance to feeding, fasting, insulin, insulin sensitivity and key metabolites in other animals, and highlight the need to understand the regulation of enzymes involved in glucose control in phocids if these animals are to be informative models of diabetes.

Effects of mass and body composition on fasting fuel utilisation in grey seal pups (Halichoerus grypus Fabricius): an experimental study using supplementary feeding

This study used supplementary feeding to test the hypothesis that fuel partitioning during the postweaning fast in grey seal pups is affected by size and composition of energy reserves at weaning, and by extra provisioning. Mass and body composition changes were measured during suckling and fasting to investigate the effect of natural differences in energy reserves at weaning on subsequent allocation of fat and protein to energy use. We fed seven pups for 5 days after weaning, to investigate the effect of increased fuel availability, and particularly protein, on fuel utilisation. After correcting for protein used during the moult, the proportional contribution of fat was 86–99% of total energy use. Pups with greater energy reserves, i.e. those that were heavier and fatter at weaning, had higher rates of fat and energy use. There was no significant relationship between adiposity at weaning and proportional contribution of fat to energy use, perhaps due to a limited sample size or range of body masses and adiposity. Supplemented individuals used energy, specifically fat, much faster and utilised proportionally less of their endogenous protein by departure than non-supplemented individuals. Fat metabolism contributed a similar percentage to daily energy use in both groups. These findings show that pups spare protein, even when energy use is dramatically increased. Pups that receive greater maternal provisioning and lay down more protein may have increased survival chances at sea. This study highlights the importance of protein reserves in first year survival of grey seal pups.